title: Herpes simplex virus 1 protein kinase Us3 phosphorylates viral envelope glycoprotein B and regulates its expression on the cell surface.
abstract: Us3 is a serine-threonine protein kinase encoded by herpes simplex virus 1 (HSV-1). As reported here, we attempted to identify the previously unreported physiological substrate of Us3 in HSV-1-infected cells. Our results were as follows. (i) Bioinformatics analysis predicted two putative Us3 phosphorylation sites in the viral envelope glycoprotein B (gB) at codons 557 to 562 (RRVSAR) and codons 884 to 889 (RRNTNY). (ii) In in vitro kinase assays, the threonine residue at position 887 (Thr-887) in the gB domain was specifically phosphorylated by Us3, while the serine residue at position 560 was not. (iii) The phosphorylation of gB Thr-887 in Vero cells infected with wild-type HSV-1 was specifically detected using an antibody that recognized phosphorylated serine or threonine residues with arginine at the -3 and -2 positions. (iv) The phosphorylation of gB Thr-887 in infected cells was dependent on the kinase activity of Us3. (v) The replacement of Thr-887 with alanine markedly upregulated the cell surface expression of gB in infected cells, whereas replacement with aspartic acid, which sometimes mimics constitutive phosphorylation, restored the wild-type phenotype. The upregulation of gB expression on the cell surface also was observed in cells infected with a recombinant HSV-1 encoding catalytically inactive Us3. These results supported the hypothesis that Us3 phosphorylates gB and downregulates the cell surface expression of gB in HSV-1-infected cells.
authors.affiliation: Division of Viral Infection, Department of Infectious Disease Control, International Research Center for Infectious Diseases, The Institute of Medical Science, The University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan.
authors.forename: Akihisa
authors.initials: A
authors.lastname: Kato
authors.forename: Jun
authors.initials: J
authors.lastname: Arii
authors.forename: Ikuo
authors.initials: I
authors.lastname: Shiratori
authors.forename: Hiroomi
authors.initials: H
authors.lastname: Akashi
authors.forename: Hisashi
authors.initials: H
authors.lastname: Arase
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Kawaguchi
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D015967:Gene Expression Regulation, Viral; D018259:Herpesvirus 1, Human / Q000502:physiology*; D006801:Humans; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D014759:Viral Envelope Proteins / Q000378:metabolism*; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D050505:Mutant Proteins; D014759:Viral Envelope Proteins; D014764:Viral Proteins; C040495:glycoprotein B, Simplexvirus; D017346:Protein Serine-Threonine Kinases; C095207:US3 protein, Human herpesvirus 1